Interpretive Summary: The mint family (Lamiaceae) provides many important commercial herb and specialty crop plants. Lamiaceae plants provide a multitude of commercially valuable secondary metabolites. Elevating CO2 levels has been employed to increase plant biomass; however, its influence on secondary metabolites in plants is unclear with many studies expressing conflictive results. In this study, we examined the influence of elevated CO2 levels on secondary products with two different Lamiaceae species with dissimilar metabolic pathways with plants grown in vivo (in soil) and in vitro (in culture media). We were able to determine that elevating CO2 and a number of other factors significantly enhanced secondary metabolite production for plants grown both in vitro and in vivo. Further, these results suggest that secondary metabolite production in vitro can exceed that occurring in vivo. Application of elevated CO2 levels to enhance secondary metabolite production has commercial and ecological significance.

Technical Abstract:
The growth (fresh weight), morphogenesis (leaves, roots, and shoots), and essential oil composition of mint (Mentha sp. L.) and thyme (Thymus vulgaris L.) plants were determined after 8 weeks under 350, 1,500, 3,000, and 30,000 uL CO2 liter**-1. Plants were grown in vitro on basal medium (BM) composed of Murashige and Skoog (MS) salts and 0.8% agar containing either 0 or 3% sucrose and employing a 180 umoles**-1 m**-2 16 hr photoperiod or in soil within a greenhouse employing natural sunlight. Ultra-high CO2 levels (i.e., >3,000 ul CO2 liter**-1) substantially increased fresh weight, leaves, shoots, and roots for all plants compared to plants grown on the same BM under ambient air (350 uL CO2 liter**-1) both in vivo and in vitro. Essential oil composition (i.e., monoterpenes, piperitonone oxide, and limonene from mint and aromatic phenol, thymol from thyme) was analyzed by CH2Cl2 extracts via gas chromatography from the shoot portion of plants grown at all CO2 levels. Higher levels of secondary compounds occurred in vitro when cultures were grown under ultra-high CO2 levels than in ambient air. Thymol, a major secondary compound in thyme plants grown on BM containing sucrose, was 317-fold higher at 10,000 uL CO2 liter **-1 than that which occurred in plants grown in soil under ambient air level with the same BM.